1. Field of the Invention
This invention relates to a method for quantitative analysis of hydrogen peroxide and a regent for use in such a quantitative analysis.
2. Description of the Background
Measuring components contained in body fluids such as blood and urine has become very important in recent years for the diagnosis of diseases, and determination of curing effects and cause of diseases.
Among various methods for measuring body fluid components heretofore developed and adopted, quantitative analyses of hydrogen peroxide are widely used and have become very important. A commonly accepted quantitative analysis of hydrogen peroxide comprises reacting a suitable oxidase with a component to be analyzed, producing hydrogen peroxide of the amount corresponding to the amount of the component to be analyzed contained in the sample, having a reagent for the analysis of hydrogen peroxide colored, and then carrying out the colorimetric analysis to determine the amount of said component. This method is utilized for the quantitative analysis of cholesterol, neutral fat, glucose, phospholipid, free fatty acid, uric acid, inorganic phosphorous compounds, pyrbic acid, L-lactic acid, amino acids, and the like, contained, for example, in blood or urine. The method is also applied to the quantitative analysis of enzymatic activities, including activities of choline esterase, .alpha.-amylase, monoamine oxidase, transaminase, or the like.
A coloring reagent generally used for the quantitative analysis of hydrogen peroxide is a combination of 4-aminoantipyrine and phenol, aniline, toluidine, anisidine, or derivatives of these compounds [for instance, Biochemistry, 6, 24-27 (1969)]. Oxidative condensates produced through such a combination of compounds, however, have a rather short maximum absorbance wavelength (.lambda.max), e.g. in the neighborhood of 500 nm, and for this reason they are liable to be affected by colored substances commonly existing in a body fluid such as hemoglobin, bilirubin, or the like. They are also largely affected by the turbidity. In addition, their sensitivity was low, so that the method was not effective for the analysis of a component which is contained in a sample at a very small amount.
The present inventors have conducted extensive studies for the development of a quantitative analysis overcoming these problems, and as a result found that the combined use of a divalent cobalt compound and a trivalent cobalt indicator can provide a highly sensitive quantitative analysis of hydrogen peroxide at a wavelength greater than 600 nm. Such a finding has led to the completion of this invention.